Systemic and immunotoxicity induced by topical application of perfluorohexane sulfonic acid (PFHxS) in a murine model.

Per- and polyfluoroalkyl substances (PFAS) are a large group of stable synthetic surfactants that are incorporated into numerous products for their water and oil resistance and have been associated with adverse health effects. The present study evaluated the systemic and immunotoxicity of sub-chronic 28- or 10-day dermal exposure of PFHxS (0.625-5% or 15.63-125 mg/kg/dose) in a murine model. Elevated levels of PFHxS were detected in the serum and urine, suggesting that absorption is occurring through the dermal route. Liver weight (% body) significantly increased and spleen weight (% body) significantly decreased with PFHxS exposure, which was supported by histopathological changes. Additionally, PFHxS significantly reduced the humoral immune response and altered immune subsets in the spleen, suggesting immunosuppression. Gene expression changes were observed in the liver, skin, and spleen with genes involved in fatty acid metabolism, necrosis, and inflammation. Immune-cell phenotyping identified significant decreases in B-cells, NK cells, and CD11b+ monocyte/macrophages in the spleen along with increases in CD4+ and CD8+ T-cells, NK cells, and neutrophils in the skin. These findings support dermal PFHxS-induced liver damage and immune suppression. Overall, data support PFHxS absorption through the skin and demonstrate immunotoxicity via this exposure route, suggesting the need for further examination.

MicroRNAs are associated with cardiac biomarkers, cardiac structure and function and incident outcomes in heart failure.

AIMS: The association between microRNAs (miRNAs) and established cardiac biomarkers is largely unknown. We aimed to measure the association between plasma miRNAs and N-terminal pro-B-type natriuretic peptide (NT-proBNP), cardiac troponin I, soluble urokinase-type plasminogen activator receptor (suPAR), and galectin-3 with cardiac structure and function and clinical outcomes. METHODS AND RESULTS: We quantified 32 plasma miRNAs using the FirePlex miRNA assay and measured biomarkers in 139 individuals with symptomatic heart failure (HF). We used principal component (PC) analysis and linear regression to evaluate the association between miRNAs and biomarkers with ventricular size and function by echocardiography and Cox modelling for the incidence of a first composite event of HF hospitalization, heart transplant, left ventricular assist device implant, or death. The mean (standard deviation) age at baseline was 64.3 (12.4) years, 33 (24%) were female, and 122 (88%) were White. A total of 45 events occurred over a median follow-up of 368 (interquartile range 234, 494) days. Baseline NT-proBNP (ß = -2.0; P = 0.001) and miRNA PC2 (ß = 2.6; P = 0.002) were associated with baseline left ventricular ejection fraction. NT-proBNP (ß = 20.6; P = 0.0004), suPAR (ß = -39.6; P = 0.005), and PC4 (ß = 21.1; P = 0.02) were associated with baseline left ventricular end-diastolic volumes. NT-proBNP [hazard ratio (HR) 1.67, 95% confidence interval (CI) 1.28-2.18, P = 0.0002], galectin-3 (HR 2.02, 95% CI 1.05-3.91, P = 0.036), PC3 (HR 1.75, 95% CI 1.23-2.49, P = 0.002), and PC4 (HR 1.67, 95% CI 1.1-2.52, P = 0.016) were independently associated with incident events. CONCLUSIONS: Biomarkers and miRNA PCs are associated with cardiac structure and function and incident cardiovascular outcomes. Combining information from miRNAs provides prognostic information beyond biomarkers in HF.

Validation of the novel GLAS algorithm as an aid in the detection of liver fibrosis and cirrhosis based on GP73, LG2m, age, and sex.

BACKGROUND: Diagnosis of liver disease at earlier stages can improve outcomes and reduce the risk of progression to malignancy. Liver biopsy is the gold standard for diagnosis of liver disease, but is invasive and sample acquisition errors are common. Serum biomarkers for liver function and fibrosis, combined with patient factors, may allow for noninvasive detection of liver disease. In this pilot study, we tested and validated the performance of an algorithm that combines GP73 and LG2m serum biomarkers with age and sex (GLAS) to differentiate between patients with liver disease and healthy individuals in two independent cohorts. METHODS: To develop the algorithm, prototype immunoassays were used to measure GP73 and LG2m in residual serum samples collected between 2003 and 2016 from patients with staged fibrosis and cirrhosis of viral or non-viral etiology (n = 260) and healthy subjects (n = 133). The performance of five predictive models using combinations of age, sex, GP73, and/or LG2m from the development cohort were tested. Residual samples from a separate cohort with liver disease (fibrosis, cirrhosis, or chronic liver disease; n = 395) and healthy subjects (n = 106) were used to validate the best performing model. RESULTS: GP73 and LG2m concentrations were higher in patients with liver disease than healthy controls and higher in those with cirrhosis than fibrosis in both the development and validation cohorts. The best performing model included both GP73 and LG2m plus age and sex (GLAS algorithm), which had an AUC of 0.92 (95% CI: 0.90-0.95), a sensitivity of 88.8%, and a specificity of 75.9%. In the validation cohort, the GLAS algorithm had an estimated an AUC of 0.93 (95% CI: 0.90-0.95), a sensitivity of 91.1%, and a specificity of 80.2%. In both cohorts, the GLAS algorithm had high predictive probability for distinguishing between patients with liver disease versus healthy controls. CONCLUSIONS: GP73 and LG2m serum biomarkers, when combined with age and sex (GLAS algorithm), showed high sensitivity and specificity for detection of liver disease in two independent cohorts. The GLAS algorithm will need to be validated and refined in larger cohorts and tested in longitudinal studies for differentiating between stable versus advancing liver disease over time.

Heterozygous GABAA receptor ß3 subunit N110D knock-in mice have epileptic spasms.

OBJECTIVE: Infantile spasms is an epileptic encephalopathy of childhood, and its pathophysiology is largely unknown. We generated a heterozygous knock-in mouse with the human infantile spasms-associated de novo mutation GABRB3 (c.A328G, p.N110D) to investigate its molecular mechanisms and to establish the Gabrb3+/N110D knock-in mouse as a model of infantile spasms syndrome. METHODS: We used electroencephalography (EEG) and video monitoring to characterize seizure types, and a suite of behavioral tests to identify neurological and behavioral impairment in Gabrb3+/N110D knock-in mice. Miniature inhibitory postsynaptic currents (mIPSCs) were recorded from layer V/VI pyramidal neurons in somatosensory cortex, and extracellular multi-unit recordings from the ventral basal nucleus of the thalamus in a horizontal thalamocortical slice were used to assess spontaneous thalamocortical oscillations. RESULTS: The infantile spasms-associated human de novo mutation GABRB3 (c.A328G, p.N110D) caused epileptic spasms early in development and multiple seizure types in adult Gabrb3+/N110D knock-in mice. Signs of neurological impairment, anxiety, hyperactivity, social impairment, and deficits in spatial learning and memory were also observed. Gabrb3+/N110D mice had reduced cortical mIPSCs and increased duration of spontaneous oscillatory firing in the somatosensory thalamocortical circuit. SIGNIFICANCE: The Gabrb3+/N110D knock-in mouse has epileptic spasms, seizures, and other neurological impairments that are consistent with infantile spasms syndrome in patients. Multiple seizure types and abnormal behaviors indicative of neurological impairment both early and late in development suggest that Gabrb3+/N110D mice can be used to study the pathophysiology of infantile spasms. Reduced cortical inhibition and increased duration of thalamocortical oscillatory firing suggest perturbations in thalamocortical circuits.

Development of exploratory algorithms to aid in risk of malignancy prediction of indeterminate pulmonary nodules.

Early detection of lung cancer allows for earlier stage treatment initiation and improved patient prognosis. This report focuses on utilization of combining patient demographic information with non-invasive biomarkers and their potential ability to predict risk of malignancy of nodules. A pilot study cohort of 141 subjects with IPNs (105 stage I cancer and 36 benign nodules) were collected by RUMC. The demographic variables of gender, age, sex, race, ethnicity, nodule size (mm), and smoking pack years, as well as the plasma levels of CA-125, SCC, CEA, HE4, ProGRP, NSE, Cyfra 21-1, hs-CRP, Ferritin, IgG, IgG1, IgG2, IgG3, IgG4, IgE, IgM, IgA, KFLC, and LFLC, were assessed for this cohort. Multivariable analyses of the previously aforementioned biomarkers and demographic variables yielded a reduced algorithm consisting of CA-125, total IgG, IgA, IgM, IgE, LFLC, nodule size, and smoking pack years with improved performance (AUC 0.82, 95 %CI 0.74-0.90) over the same analysis of the demographic variables (age, nodule size, and smoking pack years) alone (AUC 0.70, 95 %CI 0.61-0.78). This reduced algorithm of biomarkers and demographic variables may aid in assessing the risk of IPN malignancy which could be a useful stratification tool in early detection of lung cancer in high-risk subjects.

Biomarker prognostication across Universal Definition of Heart Failure stages.

AIM: The Universal Definition of Heart Failure (UDHF) provides a framework for staging risk for HF events. It is not clear whether prognostic biomarkers have different meaning across UDHF stages. We sought to evaluate performance of biomarkers to predict HF events among high-risk patients undergoing coronary and/or peripheral angiography categorized into UDHF stages. METHODS: One thousand two hundred thirty-five individuals underwent coronary and/or peripheral angiography were enrolled. Study participants were categorized into UDHF Stage A (at risk), Stage B (pre-HF), and Stage C or D (HF, including end stage) and grouped into Stage A/B and C/D. Biomarkers and clinical variables were used to develop prognostic models. Other measures examined included total HF hospitalizations. RESULTS: Over a median of 3.67 years of follow-up, 155 cardiovascular (CV) deaths occurred, and 299 patients were hospitalized with acute HF. In patients with Stage A/B, galectin-3 (HR = 1.52, P = 0.03), endothelin-1 (HR = 2.16, P = 0.001), and N-terminal pro-B-type natriuretic peptide (NT-proBNP; HR = 1.43, P < 0.001) were associated with incident CV death/HF hospitalization. In Stage C/D, NT-proBNP (HR = 1.26, P = 0.006), soluble urokinase-type plasminogen activator receptor (suPAR; HR = 1.57, P = 0.007) and high-sensitivity C-reactive protein (hs-CRP; HR = 1.15, P = 0.01) were associated with these outcomes. Higher biomarker concentrations were associated with greater total burden of HF events in Stages A/B and C/D. CONCLUSIONS: Among higher risk individuals undergoing angiographic procedures, different biomarkers improve risk stratification in different UDHF stages of HF. More precise prognostication may offer a window of opportunity to initiate targeted preventive measures.

Improving malignancy risk prediction of indeterminate pulmonary nodules with imaging features and biomarkers.

BACKGROUND: Non-invasive biomarkers are needed to improve management of indeterminate pulmonary nodules (IPNs) suspicious for lung cancer. METHODS: Protein biomarkers were quantified in serum samples from patients with 6-30 mm IPNs (n = 338). A previously derived and validated radiomic score based upon nodule shape, size, and texture was calculated from features derived from CT scans. Lung cancer prediction models incorporating biomarkers, radiomics, and clinical factors were developed. Diagnostic performance was compared to the current standard of risk estimation (Mayo). IPN risk reclassification was determined using bias-corrected clinical net reclassification index. RESULTS: Age, radiomic score, CYFRA 21-1, and CEA were identified as the strongest predictors of cancer. These models provided greater diagnostic accuracy compared to Mayo with AUCs of 0.76 (95 % CI 0.70-0.81) using logistic regression and 0.73 (0.67-0.79) using random forest methods. Random forest and logistic regression models demonstrated improved risk reclassification with median cNRI of 0.21 (Q1 0.20, Q3 0.23) and 0.21 (0.19, 0.23) compared to Mayo for malignancy. CONCLUSIONS: A combined biomarker, radiomic, and clinical risk factor model provided greater diagnostic accuracy of IPNs than Mayo. This model demonstrated a strong ability to reclassify malignant IPNs. Integrating a combined approach into the current diagnostic algorithm for IPNs could improve nodule management.

Serial cardiac biomarkers for risk stratification of patients with COVID-19.

OBJECTIVES: Several studies have demonstrated an association between elevated cardiac biomarkers and adverse outcomes in patients with COVID-19. However, the prognostic and predictive capability of a multimarker panel in a prospectively collected, diverse "all-comers" COVID-19 population has not been fully elucidated. DESIGN & METHODS: We prospectively assessed high sensitivity cardiac troponin I (hsTnI), NT-pro B-type Natriuretic Peptide (NT-proBNP), Galectin-3 (Gal-3), and procalcitonin (PCT) in 4,282 serial samples from 358 patients admitted with symptomatic, RT-PCR confirmed SARS-CoV-2 infection. Outcomes examined were 30-day in-hospital mortality and requirement for intubation within 10 days. RESULTS: Baseline hsTnI had the highest AUC for predicting 30-day mortality (0.81; 95% CI, 0.73-0.88), followed by NT-proBNP (0.80; 0.74-0.86), PCT (0.77; 0.70-0.84), and Gal-3 (0.68; 0.60-0.76). HsTnI < 3.5 ng/L at baseline identified patients at low risk for in-hospital mortality (NPV 95.9%, sensitivity 97.3%) and 10-day intubation (NPV 90.4%, sensitivity 88.5%). Continuous, log-2 increases in troponin concentration were associated with reduced survival (p < 0.001) on Kaplan-Meier curves and increased risk of 30-day mortality: HR 1.26 (1.16-1.37) in univariate and 1.19 (1.03-1.4) in multivariate models. Time-varying doubling of concentrations of hsTnI and Gal-3 were associated with increased risk of 30-day mortality (adjusted HR 1.21, 1.06-1.4, and 1.92, 1.40-2.6). CONCLUSION: HsTnI, NT-proBNP, Gal-3, and PCT are elevated at baseline in patients that have worse outcomes from COVID-19. HsTnI was the only independent predictor of 30-day mortality and intubation. Time-varying, doubling in hsTnI and Gal-3 further aided in prognostication of adverse outcomes. These results support the use of hsTnI for triaging patients with COVID-19.

Relation of High-Sensitivity Cardiac Troponin I and Obstructive Coronary Artery Disease in Patients Without Acute Myocardial Infarction.

The relation of high-sensitivity cardiac troponin I (hs-cTnI) concentration and presence of obstructive coronary artery disease (CAD) in patients without myocardial infarction (MI) is unclear. Study participants selected from patients free of MI who underwent coronary angiography with or without intervention were enrolled, and hs-cTnI measured. A gradient boosting model was implemented to build a model for detection of CAD. Cox proportional hazard regression was used to assess the association of hs-cTnI and adverse cardiovascular (CV) outcome. Among 978 study participants, 607 patients (62%) had CAD. Higher concentrations of hs-cTnI were associated with chronic kidney disease, heart failure, CAD, male gender, current tobacco use, anemia, age, and low-density lipoprotein cholesterol. History of CAD, male gender, type 2 diabetes mellitus, hs-cTnI, anemia, age, and high-density lipoprotein cholesterol were the most influential factors for detection of CAD. The gradient boosting model had an area under the curve of 0.82, accuracy of 75%, sensitivity of 88%, specificity of 52%, positive predictive value of 76%, and negative predictive value of 72% for detection of CAD. Increase in 1 log unit of hs-cTnI was significantly associated with increased risk of incident MI (hazard ratio [HR] 1.34, 95% confidence interval [CI] 1.22 to 1.47, p <0.001), CV mortality (HR 1. 24, 95% CI 1.11 to 1.39, p <0.001), and composite of incident MI or CV mortality (HR 1.29, 95% CI 1.20 to 1.40, p <0.001). In conclusion, among patients without acute MI and CAD, higher concentrations of hs-cTnI were associated with the presence of CAD and linked to increased risk of future CV events. ClinicalTrials.gov Identifier: NCT00842868.

Altered Channel Conductance States and Gating of GABAA Receptors by a Pore Mutation Linked to Dravet Syndrome.

We identified a de novo missense mutation, P302L, in the γ-aminobutyric acid type A (GABAA) receptor γ2 subunit gene GABRG2 in a patient with Dravet syndrome using targeted next-generation sequencing. The mutation was in the cytoplasmic portion of the transmembrane segment M2 of the γ2 subunit that faces the pore lumen. GABAA receptor α1 and ß3 subunits were coexpressed with wild-type (wt) γ2L or mutant γ2L(P302L) subunits in HEK 293T cells and cultured mouse cortical neurons. We measured currents using whole-cell and single-channel patch clamp techniques, surface and total expression levels using surface biotinylation and Western blotting, and potential structural perturbations in mutant GABAA receptors using structural modeling. The γ2(P302L) subunit mutation produced an ∼90% reduction of whole-cell current by increasing macroscopic desensitization and reducing GABA potency, which resulted in a profound reduction of GABAA receptor-mediated miniature IPSCs (mIPSCs). The conductance of the receptor channel was reduced to 24% of control conductance by shifting the relative contribution of the conductance states from high- to low-conductance levels with only slight changes in receptor surface expression. Structural modeling of the GABAA receptor in the closed, open, and desensitized states showed that the mutation was positioned to slow activation, enhance desensitization, and shift channels to a low-conductance state by reshaping the hour-glass-like pore cavity during transitions between closed, open, and desensitized states. Our study revealed a novel γ2 subunit missense mutation (P302L) that has a novel pathogenic mechanism to cause defects in the conductance and gating of GABAA receptors, which results in hyperexcitability and contributes to the pathogenesis of the genetic epilepsy Dravet syndrome.

Correction: Deleterious Rare Variants Reveal Risk for Loss of GABAA Receptor Function in Patients with Genetic Epilepsy and in the General Population.

[This corrects the article DOI: 10.1371/journal.pone.0162883.].

Deleterious Rare Variants Reveal Risk for Loss of GABAA Receptor Function in Patients with Genetic Epilepsy and in the General Population.

Genetic epilepsies (GEs) account for approximately 50% of all seizure disorders, and familial forms include mutations in single GABAA receptor subunit genes (GABRs). In 144 sporadic GE cases (GECs), exome sequencing of 237 ion channel genes identified 520 GABR variants. Among these variants, 33 rare variants in 11 GABR genes were present in 24 GECs. To assess functional risk of variants in GECs, we selected 8 variants found in GABRA, 3 in GABRB, and 3 in GABRG and compared them to 18 variants found in the general population for GABRA1 (n = 9), GABRB3 (n = 7), and GABRG2 (n = 2). To identify deleterious variants and gain insight into structure-function relationships, we studied the gating properties, surface expression and structural perturbations of the 32 variants. Significant reduction of GABAA receptor function was strongly associated with variants scored as deleterious and mapped within the N-terminal and transmembrane domains. In addition, 12 out of 17 variants mapped along the ß+/α- GABA binding interface, were associated with reduction in channel gating and were predicted to cause structural rearrangements of the receptor by in silico simulations. Missense or nonsense mutations of GABRA1, GABRB3 and GABRG2 primarily impair subunit biogenesis. In contrast, GABR variants affected receptor function by impairing gating, suggesting that different mechanisms are operating in GABR epilepsy susceptibility variants and disease-causing mutations. The functional impact of single GABR variants found in individuals with sporadic GEs warrants the use of molecular diagnosis and will ultimately improve the treatment of genetic epilepsies by using a personalized approach.

Loss of CCR5 results in glucose intolerance in diet-induced obese mice.

Macrophage and T cell infiltration into metabolic tissues contributes to obesity-associated inflammation and insulin resistance (IR). C-C chemokine receptor 5 (CCR5), expressed on macrophages and T cells, plays a critical role in the recruitment and activation of proinflammatory M1 and TH1 immune cells to tissues and is elevated in adipose tissue (AT) and liver of obese humans and mice. Thus, we hypothesized that deficiency of CCR5 would protect against diet-induced inflammation and IR. CCR5-deficient (CCR5(-/-)) mice and C57BL/6 (WT) controls were fed 10% low-fat (LF) or 60% high-fat (HF) diets for 16 wk. HF feeding increased adiposity, blood glucose, and plasma insulin levels equally in both genotypes. Opposing our hypothesis, HF-fed CCR5(-/-) mice were significantly more glucose intolerant than WT mice. In AT, there was a significant reduction in the M1-associated gene CD11c, whereas M2 associated genes were not different between genotypes. In addition, HF feeding caused a twofold increase in CD4(+) T cells in the AT of CCR5(-/-) compared with WT mice. In liver and muscle, no differences in immune cell infiltration or inflammatory cytokine expression were detected. However, in AT and muscle, there was a mild reduction in insulin-induced phosphorylation of AKT and IRß in CCR5(-/-) compared with WT mice. These findings suggest that whereas CCR5 plays a minor role in regulating immune cell infiltration and inflammation in metabolic tissues, deficiency of CCR5 impairs systemic glucose tolerance as well as AT and muscle insulin signaling.

Irritancy and allergic responses induced by exposure to the indoor air chemical 4-oxopentanal.

Over the last two decades, there has been an increasing awareness regarding the potential impact of indoor air pollution on human health. People working in an indoor environment often experience symptoms such as eye, nose, and throat irritation. Investigations into these complaints have ascribed the effects, in part, to compounds emitted from building materials, cleaning/consumer products, and indoor chemistry. One suspect indoor air contaminant that has been identified is the dicarbonyl 4-oxopentanal (4-OPA). 4-OPA is generated through the ozonolysis of squalene and several high-volume production compounds that are commonly found indoors. Following preliminary workplace sampling that identified the presence of 4-OPA, these studies examined the inflammatory and allergic responses to 4-OPA following both dermal and pulmonary exposure using a murine model. 4-OPA was tested in a combined local lymph node assay and identified to be an irritant and sensitizer. A Th1-mediated hypersensitivity response was supported by a positive response in the mouse ear swelling test. Pulmonary exposure to 4-OPA caused a significant elevation in nonspecific airway hyperreactivity, increased numbers of lung-associated lymphocytes and neutrophils, and increased interferon-γ production by lung-associated lymph nodes. These results suggest that both dermal and pulmonary exposure to 4-OPA may elicit irritant and allergic responses and may help to explain some of the adverse health effects associated with poor indoor air quality.

The identification of a sensitizing component used in the manufacturing of an ink ribbon.

Skin diseases including dermatitis constitute ≈ 30% of all occupational illnesses, with a high incidence in the printing industry. An outbreak of contact dermatitis among employees at an ink ribbon manufacturing plant was investigated by scientists from the National Institute for Occupational Safety and Health (NIOSH). Employees in the process areas of the plant were exposed to numerous chemicals and many had experienced skin rashes, especially after the introduction of a new ink ribbon product. To identify the causative agent(s) of the occupational dermatitis, the murine local lymph node assay (LLNA) was used to identify the potential of the chemicals used in the manufacture of the ink ribbon to induce allergic contact dermatitis. Follow-up patch testing with the suspected allergens was conducted on exposed employees. Polyvinyl butyral, a chemical component used in the manufacture of the ink ribbon in question and other products, tested positive in the LLNA, with an EC3 of 3.6%, which identifies it as a potential sensitizer; however, no employees tested positive to this chemical during skin patch testing. This finding has implications beyond those described in this report because of occupational exposure to polyvinyl butyral outside of the printing industry.

Evaluation of furfuryl alcohol sensitization potential following dermal and pulmonary exposure: enhancement of airway responsiveness.

Furfuryl alcohol is considered by the U.S. Environmental Protection Agency to be a high volume production chemical, with over 1 million pounds produced annually. Due to its high production volume and its numerous industrial and consumer uses, there is considerable potential for work-related exposure, as well as exposure to the general population, through pulmonary, oral, and dermal routes of exposure. Human exposure data report a high incidence of asthma in foundry mold workers exposed to furan resins, suggesting potential immunologic effects. Although furfuryl alcohol was nominated and evaluated for its carcinogenic potential by the National Toxicology Program, studies evaluating its immunotoxicity are lacking. The studies presented here evaluated the immunotoxic potential of furfuryl alcohol following exposure by the dermal and pulmonary routes using a murine model. When tested in a combined irritancy local lymph node assay, furfuryl alcohol was identified to be an irritant and mild sensitizer (EC3 = 25.6%). Pulmonary exposure to 2% furfuryl alcohol resulted in enhanced airway hyperreactivity, eosinophilic infiltration into the lungs, and enhanced cytokine production (IL-4, IL-5, and interferon-γ) by ex vivo stimulated lung-associated draining lymphoid cells. Airway hyperreactivity and eosinophilic lung infiltration were augmented by prior dermal exposure to furfuryl alcohol. These results suggest that furfuryl alcohol may play a role in the development of allergic airway disease and encourage the need for additional investigation.

Allergic Potential and Immunotoxicity Induced by Topical Application of 1-Chloro-4-(Trifluoromethyl)Benzene (PCBTF) in a Murine Model.

The purpose of the studies in this paper was to evaluate the allergic potential, immunotoxicity, and irritancy of the occupationally relevant chemical, 1-chloro-4-(trifluoromethyl)benzene, also known as parachlorobenzotrifluoride (PCBTF), following dermal exposure in a murine model. Evaluation of the sensitization potential, conducted using the local lymph node assay (LLNA) at concentrations ranging from 50% to 100%, identified a dose-dependent increase in lymphocyte proliferation with a calculated EC3 value of 53.1%. While no elevations in total or specific IgE were observed after exposure to any concentration of the chemical, significant increases in IFN-γ protein production by stimulated draining lymphoid cells were observed, indicating a T-cell-mediated response. Dermal exposure to PCBTF was not found to alter the immune response to a T-cell-dependant antigen. These results demonstrate that PCBTF has the potential to induce allergic sensitization following dermal exposure and based on LLNA results would be classified as a weak sensitizer.

Evaluation of dicarbonyls generated in a simulated indoor air environment using an in vitro exposure system.

Over the last two decades, there has been increasing awareness regarding the potential impact of indoor air pollution on health. Exposure to volatile organic compounds (VOCs) or oxygenated organic compounds formed from indoor chemistry has been suggested to contribute to adverse health effects. These studies use an in vitro monitoring system called VitroCell, to assess chemicals found in the indoor air environment. The structurally similar dicarbonyls diacetyl, 4-oxopentanal (4-OPA), glyoxal, glutaraldehyde, and methyl glyoxal were selected for use in this system. The VitroCell module was used to determine whether these dicarbonyls were capable of inducing inflammatory cytokine expression by exposed pulmonary epithelial cells (A549). Increases in the relative fold change in messenger RNA expression of the inflammatory mediators, interleukin (IL)-6, IL-8, granulocyte-macrophage colony-stimulating factor (GM-CSF), and tumor necrosis factor alpha (TNF-alpha) were identified following exposure to diacetyl, 4-OPA, glyoxal, glutaraldehyde, and methyl glyoxal when compared to a clean air control. Consistent results were observed when the protein levels of these cytokines were analyzed. Exposure to 4-OPA significantly elevated IL-8, IL-6, GM-CSF, and TNF-alpha while glutaraldehyde caused significant elevations in IL-6, IL-8, and TNF-alpha. IL-6 and IL-8 were also significantly elevated after exposure to diacetyl, glyoxal, and methyl glyoxal. These studies suggest that exposure to structurally similar oxygenated reaction products may be contributing to some of the health effects associated with indoor environments and may provide an in vitro method for identification and characterization of these potential hazards.

Irritancy and allergic responses induced by topical application of ortho-phthalaldehyde.

Although ortho-phthalaldehyde (OPA) has been suggested as an alternative to glutaraldehyde for the sterilization and disinfection of hospital equipment, the toxicity has not been thoroughly investigated. The purpose of these studies was to evaluate the irritancy and sensitization potential of OPA. The EpiDerm Skin Irritation Test was used to evaluate in vitro irritancy potential of OPA and glutaraldehyde. Treatment with 0.4125 and 0.55% OPA induced irritation, while glutaraldehyde exposure at these concentrations did not. Consistent with the in vitro results, OPA induced irritancy, evaluated by ear swelling, when mice were treated with 0.75%. Initial evaluation of the sensitization potential was conducted using the local lymph node assay at concentrations ranging from 0.005 to 0.75%. A concentration-dependent increase in lymphocyte proliferation was observed with a calculated EC3 value of 0.051% compared to that of 0.089%, previously determined for glutaraldehyde. Immunoglobulin (Ig) E-inducing potential was evaluated by phenotypic analysis of draining lymph node (DLN) cells and measurement of total and specific serum IgE levels. The 0.1 and 0.75% exposed groups yielded significant increases in the IgE+B220+ cell population in the lymph nodes while the 0.75% treated group demonstrated significant increases in total IgE, OPA-specific IgE, and OPA-specific IgG(1). In addition, significant increases in interleukin-4 messenger RNA and protein expression in the DLNs were observed in OPA-treated groups. The results demonstrate the dermal irritancy and allergic potential of OPA and raise concern about the proposed/intended use of OPA as a safe alternative to glutaraldehyde.

Evaluation of irritancy and sensitization potential of metalworking fluid mixtures and components.

There are approximately 1.2 million workers exposed to metalworking fluids (MWF), which are used to reduce the heat and friction associated with industrial machining and grinding operations. Irritancy and sensitization potential of 9 National Toxicology Program (NTP) nominated MWFs (TRIM 229, TRIM VX, TRIM SC210, CIMTECH 310, CIMPERIAL 1070, CIMSTAR 3800, SYNTILO 1023, SUPEREDGE 6768, and CLEAREDGE 6584) were examined in a combined local lymph node assay (LLNA). BALB/c mice were dermally exposed to each MWF at concentrations up to 50%. Significant irritation was observed after dermal exposure to all MWFs except CIMTECH 310 and SYNTILO 1023. Of the 9 MWFs, 6 induced greater than a 3-fold increase in lymphocyte proliferation and 7 tested positive in the irritancy assay. TRIM VX yielded the lowest EC3 value (6.9%) with respect to lymphocyte proliferation. Chemical components of TRIM VX identified using HPLC were screened for sensitization potential using structural activity relationship (SAR) modeling and the LLNA. TOPKAT predicted triethanolamine (TEA) as a sensitizer while Derek for Windows predicted only 4-chloro-3-methylphenol (CMP) to be positive for sensitization. When tested in the LLNA only CMP (EC3 = 11.6%) and oleic acid (OA) (EC3 = 29.7%) were identified as sensitizers. Exposure to all tested TRIM VX components resulted in statistically significant irritation. An additive proliferative response was observed when mixtures of the two identified sensitizing TRIM VX components, OA and CMP, were tested in the LLNA. This is one explanation of why the EC3 value of TRIM VX, with respect to lymphocyte proliferation, is lower than those assigned to its sensitizing components.